Roth was educated at the universities of Tübingen and Berlin. He continued his studies in Paris and London, and in 1848 was appointed as extraordinary professor of Oriental languages in Tübingen University, becoming a full professor and principal librarian in 1856.
His chief work is the monumental Sanskrit Wörterbuch (Sanskrit dictionary, 7 vols., Saint Petersburg, 1853–1895), compiled in collaboration with Otto von Böhtlingk and published by the Saint Petersburg Academy of Sciences. He edited Yaska's Nirukta (1852) and, with Whitney, the Atharva Veda (1856–1857).
The original works of Roth include: Zur Litteratur und Geschichte des Veda (On the literature and history of the Veda, 1846), a ground-breaking work on Vedic scholarship and research; Ueber den Mythus von den fünf Menschengeschlechtern ("On the myth of the five races of humans", 1860); Ueber die Vorstellung vom Schicksal in der indischen Sprachweisheit ("On the representation of fate in Indian wisdom literature", 1866); Der Atharva-Veda in Kaschmir (1875); and Ueber Yaçna 31 (1876).